The binding of carcinogens and antitumor drug analogs in the situation of high DNA base pair-to-bound carcinogen/drug ratios is investigated. With antitumor drug analogs the role of side chains in the observed cooperativity and the effects of the side chains on DNA structure and functions are examined. Analogs of antitumor drugs consisting of either the chromophore 9-aminoacridine or 1,4-diamino-9,10-anthracenedione substituted on the amino moieties with various alkyl amine side chains are synthesized and the binding of these molecules to DNAs monitored by spectrophotometric titrations or phase partition techniques. Binding of the different molecules will be analyzed and compared in terms of the relative amounts of cooperativity in drug binding produced by the different side chains. 1HNMR and CD studies of the effects of different side chains on oligonucleotide structure and RNA polymerase activity studies with the different derivatives give correlations between cooperativity and the side chains. These studies provide insight into which drug structural features affect the DNA and are, hence, important for drug activity. Using the covalently binding metabolites (radioactively labeled) of the carcinogens acetylaminofluorene and 4-nitroquinoline-1-oxide and restriction endonucleases, the small number of "high affinity" carcinogen binding sites on the plasmid pBR 322 are located. Then, fragments containing these binding sites will be isolated for (1) studies of the equilibrium binding of carcinogens to these fragments; (2) sequencing of the fragments to find the exact carcinogen binding sites; (3) studies, using NMR, CD, DNA enzymes, of the effects of carcinogen binding into these sites on DNA structures and functions. From such studies, further information on how carcinogens act on DNA to lead to DNA malfunctions may be gleaned.